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Researchers Observe Changes in Fish Cell Proteomes to Study Water Contamination


While whole fish are usually used to test aquatic environments for potentially lethal contaminants, a group of Canadian researchers have devised another method of testing the waters by monitoring the proteomic changes of fish cell lines.

"The 96-hour lethality test for trout is costly and time consuming," said Lucy Lee, a professor of biology at Wilfred Laurier University in Waterloo, Ontario, who cultivates and maintains numerous fish cell lines. "With protein assays we can see differences within 12 hours. Depending on the protein profile you see, you can say, 'These cells have been exposed to such and such a contaminant.'"

While proteomics has been used to study mammalian cell lines for many years, it has not been applied very much to the fish community, Lee noted.

"Fish people are actually lagging far behind mammalian people," said Lee. "But the [proteomic] technology should be applied to all types of areas."

As a kind of proof of principle to demonstrate that proteomics can be used to study fish cell lines, Lee and her undergraduate student Sarah Wagg used 2D gels to show that protein spot patterns can be used to distinguish different fish cell lines from each other.

The study, published in the Sept. 30 issue of Proteomics, shows that while different cells within the cell line may have slightly different 2D gel protein spot profiles, when the gels are "averaged out" they exhibit a profile that is distinctly different from the averaged-out spot profiles of other cell lines.

"When you look at eel cells, they are definitely out of comparison with the trout, and even if the cell lines are from the same organism, you can tell a liver cell from trout from a gill cell from the same trout," said Lee.

"Fish people are actually lagging far behind mammalian people. But the [proteomic] technology should be applied to all types of areas."

The ability to distinguish proteomic spot profiles of different cell lines may help determine if a cell line has become contaminated with another cell line, Lee noted.

Currently, PCR is usually used to test for cell-line contamination, but PCR can not distinguish between different organs of the same organism, said Lee. While 2D proteomic gels are not necessarily easier to do than PCR, they are at least simple enough to be done by undergraduate students, and they can distinguish between different organs within the same fish, she added.

Lee is currently working on publishing work she has done that shows that proteomic changes can be used to monitor for environmental contaminants in water. She is also working to distinguish protein profiles of fish cell lines that have been exposed to pathogens such as marine amoebas.

"Marine amoebas are normally found to be harmless, but they could become virulent," said Lee. "We're looking at studying the mechanisms of virulence."

One of the main reasons for developing cell lines of any kind is to study viruses, Lee pointed out.

"They didn't want to infect the whole organism all the time, so they developed cell lines for growing viruses," said Lee.

Within the fish world, quite a lot of viruses have emerged, such as hemorrhagic viruses and noda viruses, but little is known about the viruses because they are not well studied, said Lee.

"It's not known if they are a threat to humans. Some viruses could jump species, but it's not known now if [these viruses] can," she said.

Though fishing is a big industry, and demand for fish is high, diseases in fish are barely being studied, Lee said.

"How many fish vets are there out there? Not many," said Lee.

One problem with studying any type of contagious virus, including fish viruses, is that experiments must be done within secure laboratories, such as Biosafety Level 2 or BSL 3 laboratories, Lee noted. The laboratories are expensive to maintain.

Lee has published papers on five of her fish cell lines: a liver, a gill, and two pituitary cell lines in rainbow trout, and a skin cell line in goldfish. She is currently working on publishing papers on another five cell lines, and on creating new fish cell lines.

"A lot of fish biologists want to have a brain cell line to study neurotoxicity, so I'm working on developing one of those," said Lee.

Though Lee may eventually use mass spectrometry to identify a protein from a particular protein spot of interest, so far she is relying solely on 2D gels for proteomic work because her laboratory does not have the money to purchase a mass spec, nor the personnel to run it.

"Eventually if we find something that's really interesting, we may use the mass spectrometer across the street at the University of Waterloo to identify it," said Lee. "But you're talking to someone who has a very small lab. We're doing all work at a minimum budget. Mass spec is way too expensive for us."

Lee said her primary interest for her lab is in developing fish cell lines so that other people can use the cells to study toxicology, immunology, or other fields of interest. She gets a lot of requests for the fish cell lines because there aren't too many people who produce them.

As a secondary interest, Lee develops applications for her cell lines, such as studying the proteomic profiles of cell lines that are grown in contaminated waters.

Asked if she would consider commercializing her fish cell lines, Lee said, "I guess I could if the demand is high. But right now, we're just giving them away."

— Tien-Shun Lee ([email protected])

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